The cIAP ubiquitin ligases sustain type 3 γδ T cells and ILC during aging to promote barrier immunity.

Early-life cues shape the immune system during adulthood. However, early-life signaling pathways and their temporal functions are not well understood. Herein, we demonstrate that the cellular inhibitor of apoptosis proteins 1 and 2 (cIAP1/2), which are E3 ubiquitin ligases, sustain interleukin (IL)-17-producing γ δ T cells (γδT17) and group 3 innate lymphoid cells (ILC3) during late neonatal and prepubescent life. We show that cell-intrinsic deficiency of cIAP1/2 at 3-4 wk of life leads to downregulation of the transcription factors cMAF and RORγt and failure to enter the cell cycle, followed by progressive loss of γδT17 cells and ILC3 during aging. Mice deficient in cIAP1/2 have severely reduced γδT17 cells and ILC3, present with suboptimal γδT17 responses in the skin, lack intestinal isolated lymphoid follicles, and cannot control intestinal bacterial infection. Mechanistically, these effects appear to be dependent on overt activation of the non-canonical NF-κB pathway. Our data identify cIAP1/2 as early-life molecular switches that establish effective type 3 immunity during aging.

The porcine large intestine contains developmentally distinct submucosal lymphoid clusters and mucosal isolated lymphoid follicles.

Gut-associated lymphoid tissues (GALT) serve as key priming sites for intestinal adaptive immune responses. Most of our understanding of GALT function and development arises from studies in mice. However, the diversity, structure and cellular composition of GALT differs markedly between mammalian species and the developmental window in which distinct GALT structures develop in large mammals remains poorly understood. Given the importance of pigs as models of human disease, as well as their role in livestock production, we adapted a recently developed protocol for the isolation of human GALT to assess the diversity, development and immune composition of large intestinal GALT in neonatal and adult pigs. We demonstrate that the large intestine of adult pigs contains two major GALT types; multifollicular submucosal GALT that we term submucosal lymphoid clusters (SLC) which develop prenatally, and as yet undescribed mucosal isolated lymphoid follicles (M-ILF), which arise after birth. Using confocal laser microscopy and flow cytometry, we additionally assess the microanatomy and lymphocyte composition of SLC and M-ILF, compare them to jejunal Peyer's patches (PP), and describe the maturation of these structures. Collectively, our results provide a deeper understanding of the diversity and development of GALT within the porcine large intestine.

Human gut-associated lymphoid tissues (GALT); diversity, structure, and function.

Gut-associated lymphoid tissues (GALT) are the key antigen sampling and adaptive immune inductive sites within the intestinal wall. Human GALT includes the multi-follicular Peyer's patches of the ileum, the vermiform appendix, and the numerous isolated lymphoid follicles (ILF) which are distributed along the length of the intestine. Our current understanding of GALT diversity and function derives primarily from studies in mice, and the relevance of many of these findings to human GALT remains unclear. Here we review our current understanding of human GALT diversity, structure, and composition as well as their potential for regulating intestinal immune responses during homeostasis and inflammatory bowel disease (IBD). Finally, we outline some key remaining questions regarding human GALT, the answers to which will advance our understanding of intestinal immune responses and provide potential opportunities to improve the treatment of intestinal diseases.

Identification, isolation and analysis of human gut-associated lymphoid tissues.

Gut-associated lymphoid tissues (GALTs) comprise key intestinal immune inductive sites, including the Peyer's patches of the small intestine and different types of isolated lymphoid follicle (ILF) found along the length of the gut. Our understanding of human GALT is limited due to a lack of protocols for their isolation. Here we describe a technique that, uniquely among intestinal cell isolation protocols, allows identification and isolation of all human GALT, as well as GALT-free intestinal lamina propria (LP). The technique involves the mechanical separation of intestinal mucosa from the submucosa, allowing the identification and isolation of submucosal ILF (SM-ILF), LP-embedded mucosal ILF (M-ILF) and LP free of contaminating lymphoid tissue. Individual SM-ILF, M-ILF and Peyer's patch follicles can be subsequently digested for downstream cellular and molecular characterization. The technique, which takes 4-10 h, will be useful for researchers interested in intestinal immune development and function in health and disease.

Immune Profiling of Human Gut-Associated Lymphoid Tissue Identifies a Role for Isolated Lymphoid Follicles in Priming of Region-Specific Immunity.

The intestine contains some of the most diverse and complex immune compartments in the body. Here we describe a method for isolating human gut-associated lymphoid tissues (GALTs) that allows unprecedented profiling of the adaptive immune system in submucosal and mucosal isolated lymphoid follicles (SM-ILFs and M-ILFs, respectively) as well as in GALT-free intestinal lamina propria (LP). SM-ILF and M-ILF showed distinct patterns of distribution along the length of the intestine, were linked to the systemic circulation through MAdCAM-1+ high endothelial venules and efferent lymphatics, and had immune profiles consistent with immune-inductive sites. IgA sequencing analysis indicated that human ILFs are sites where intestinal adaptive immune responses are initiated in an anatomically restricted manner. Our findings position ILFs as key inductive hubs for regional immunity in the human intestine, and the methods presented will allow future assessment of these compartments in health and disease.